Problems with balance are a leading cause of injury and even death in elderly populations. Degeneration of otoconia is thought to contribute significantly to balance disorders and to the displacement or ectopic formation of otoconia that occur in patients suffering from benign paroxysmal vertigo (BPV). Despite the prevalence of balance disorders, little is known about the molecular mechanisms regulating the development and pathology of the vestibular mechanosensory apparatus. This proposal aims to advance our understanding of the molecular mechanisms that regulate the development of otoconia by investigating the biological and biochemical activities of a newly discovered gene family, the Otopetrins. This knowledge will provide insight into pathologic mechanisms that could contribute to otoconial degeneration. Tilted mice have a severe balance disorder due to the congenital absence of otoconia. By positional cloning we identified mutations in Otopetrin 1 (Otop1) as the genetic etiology of the tilted mouse phenotype. Biochemical studies using a ratiometric calcium assay showed that overexpression of Otop1 has three prominent consequences: 1) nonspecific depletion of endoplasmic reticulum calcium stores, 2) specific inhibition of the purinergic receptor P2Y2, and 3) initiation of a novel influx of extracellular calcium in response to ATP-like nucleotides. The long-term objective of this research program is to understand the biochemical, developmental, and physiological functions of the Otopetrin gene family. A comprehensive understanding of Otopetrin function is essential to understand mechanisms regulating otoconial morphogenesis, maintenance and repair. This work is also likely to uncover additional roles for Otopetrin genes in the development and physiology of other tissues and organs, which in turn will aide in our understanding of otoconial development and physiology. In this proposal, we will: 1) Identify functional domains within Otop1 and determine the extent of conserved biochemical function among the three known Otopetrin proteins; 2) Identify the mechanism by which mutations in tilted (tlt), mergulhador (mlh) and inner ear defect (ied) mice alter the biochemical function of Otop1; 3) Determine whether wild type or mutant Otopetrins can modulate calcium in vestibular epithelial cells; 4) Identify developmental and physiological functions of Otop1 and Ootp2 in vivo and test the hypothesis that Otopetrins have redundant function in vivo. [unreadable] [unreadable] [unreadable]